TREATMENT of primary cultures of rat cerebral cortical astrocytes with extracellular ATP caused a 3− to 4-fold stimulation of mitogen-activated protein (MAP) kinase activity. Studies with agonists and antagonists of P1 and P2 purinergic receptors indicated that this activation is mediated by ATP/P2 purinergic receptors rather than adenosine/P1 purinergic receptors. Increased cAMP levels did not significantly inhibit the activation of MAP kinase by ATP but did inhibit the stimulation of MAP kinase by basic fibroblast growth factor, a polypeptide growth factor that activates the Ras/Raf kinase signaling pathway. These data indicate that ATP/P2 purinergic receptors are coupled to MAP kinase by a signal transduction pathway that is independent of the Raf kinase pathway.